Phenotypes and Functional Effects Caused by Various Viral RNA Silencing Suppressors in Transgenic Nicotiana benthamiana and N. tabacum

Abstract

RNA silencing suppressor genes derived from six virus genera were transformed into Nicotiana benthamiana and N. tabacum plants. These suppressors were P1 of Rice yellow mottle virus (RYMV), P1 of Cocksfoot mottle virus, P19 of Tomato bushy stunt virus, P25 of Potato virus X, HcPro of Potato virus Y (strain N), 2b of Cucumber mosaic virus (strain Kin), and AC2 of African cassava mosaic virus (ACMV). HcPro caused the most severe phenotypes in both Nicotiana spp. AC2 also produced severe effects in N. tabacum but a much milder phenotype in N. benthamiana, although both HcPro and AC2 affected the leaf tissues of the two Nicotiana spp. in similar ways, causing hyperplasia and hypoplasia, respectively. P1-RYMV caused high lethality in the N. benthamiana plants but only mild effects in the N. tabacum plants. Phenotypic alterations produced by the other transgenes were minor in both species. Interestingly, the suppressors had very different effects on crucifer-infecting Tobamovirus (crTMV) infections. AC2 enhanced both spread and brightness of the crTMV-green fluorescent protein (GFP) lesions, whereas 2b and both P1 suppressors enhanced spread but not brightness of these lesions. P19 promoted spread of the infection into new foci within the infiltrated leaf, whereas HcPro and P25 suppressed the spread of crTMV-GFP lesions.

Structural basis for RNA‐silencing suppression by Tomato aspermy virus protein 2b

“…In these transgenic plants, GFP silencing can be triggered by transient GFP expression by Agrobacterium infiltration ( Johansen & Carrington, 2001), whereas RNA-silencing suppressors, which initiate post-transcriptional gene silencing, can rescue transient expression of GFP in the infiltrated regions. Our results show that transient expression of TAV2b suppressed GFP silencing and rescued the expression of GFP (Fig 3, lower left spot on each leaf), similar to suppression by the P19 proteins (Fig 3, upper right spot on each leaf; Siddiqui et al, 2008). As a negative control, GFP protein expression was almost completely suppressed when the GFP transgene was infiltrated alone (Fig 3, upper left spot on each leaf).…”

Oxylipin Biosynthesis Genes Positively Regulate Programmed Cell Death during Compatible Infections with the Synergistic Pair Potato Virus X-Potato Virus Y and Tomato Spotted Wilt Virus

“…It has been suggested that symptoms in potyvirus-infected plants are associated at least to some extent with the RNA-silencing suppressor activity of the HC-Pro protein that interferes with endogenous microRNA (miRNA) accumulation, causes misregulation of the expression of several miRNA-regulated transcription factors, and produces developmental defects in transgenic plants (66). Similarly, transgenic expression of the silencing suppressor protein of PVX, P25, produced developmental alterations in N. benthamiana (67). Thus, PVX-PVY-associated synergism could partly be caused by the combined effect of both suppressors on several RNA-regulated networks.…”

Effects of viral silencing suppressors on tobacco ringspot virus infection in two Nicotiana species

“…The P25 transgene also prevented recovery of N. benthamiana from systemic TRSV infection, but it did not allow the systemic infection of TRSV in the transgenic N. tabacum host at elevated temperatures. This failure in this Nicotiana species was probably due to the low expression level of the transgene in this line (Siddiqui et al, 2008). On the other hand, it is conceivable that the P25 protein function itself is temperature-sensitive, as indicated by the results of Close (1964) and Xie et al (2001).…”